Labeled oligonucleotides are being used extensively as genetic probes because they are relatively easy to make and, under ideal conditions, can discriminate by hybridization between gene sequences that differ by a single base pair. The recent proliferation of sequences for the neutralizing gene of different strains of rotaviruses encouraged us to prepare oligonucleotide probes to a series of areas common to all serotypes of rotavirus as well as to areas demonstrating great serotypic diversity. We have tested these probes for their sensitivity and specificity in detecting rotavirus in stool specimens and in serotyping those specimens found to be positive. We have also compared the sensitivity of short oligonucleotide probes with larger single stranded RNA transcripts prepared according to procedures developed previously in this laboratory. Radiolabeled oligonucleotide probes appear to be about as sensitive as the ELISA test in detecting rotavirus in stool specimens and are significantly less sensitive than ssRNA transcript probes. The level of sensitivity and specificity are reduced when biotinylated oligonucleotide probes are used. Serotyping will be examined by hybridizing oligo probes to Northern blots of viral RNA since neither specificity nor sensitivity were adequate using dotted stool RNAs.